The proposed research concerns proton transport by lysosomes. This transport is necessary because proteins continually entering lysosomes are at neutral pH but lysosomal proteases require an acid pH to be active. Proton transport will be measured as uptake of radioactive methylamine by rat liver lysosomal membrane vesicles resealed in the absence and presence of lysosomal soluble proteins. The vesicle preparations will facilitate determination of the relative roles of the an ATP-driven proton pump, Donnan equilibrium and membrane potential in establishing and maintaining the pH gradient vital for lysosome function. In the second phase of the proposed research, the protein responsible for acidification of lysosomes will be isolated. If, as expected, a proton pump is of major importance, ATPase activity and acidification of resealed lysosomal membranes will be used to assay functional protein during purification. If the Donnan effect is the major Proton motive force, then a highly anionic macromolecule perhaps associated with the cell surface may be obligatorily internalized each time protein destined for degradation enters the lysosome. The significance of the proposed research is apparent from the magnitude of protein degradation, 50% of liver protein and about 10% of plasma proteins daily. Disorders in which protein degradation is aberrant, for example cardiac hypertrophy and muscular dystrophy, indicate the essential nature of protein turnover and of reutilization of components. The proposed research will help elucidate basic aspects of this important lysosomal process.